1. Field of the Invention
The United States Government has rights in this invention pursuant to contract no. DE-AC05-840R2 1400 between the United States Department of Energy and Martin Marietta Energy Systems, Inc., and funded by the Office of Health and Environmental Research.
The present invention relates to chemical sensors and biosensors, and more particularly, to a sensor that can selectively recognize and transduce the change in metal ion concentration into a measurable optical signal.
2. Description of the Related Art
Recently there has been an increasing interest in the development of practical, sensitive and miniaturized probe systems for use in monitoring metal ions such as Ca.sup.2+, in biological media, living cells, and environmental samples. With recent advances in integrated waveguide technology and advanced photometric detection systems, fiberoptic sensors are most appropriate devices for a wide variety of monitoring applications in biological, environmental and process control applications because they shield the analytical process from the sample environment, and allow remote and in situ analysis of minute sample volumes. Fluorescence based sensors are especially useful due to their sensitivity, achieving attomole (10.sup.-8 mole) limits of detection. Such sensors use probes that have chemical reagents or bioreceptors (such as an antibody) chemically bound to optical fibers or physically entrapped in sensing microcavities containing liquid reagents or gels attached to the distal end or to the cladding of the optical fiber. Direct attachment allows fast response time since sensor response depends on the mass transfer rate of the analyte to the immobilized reagent. In some cases, gels may be saturated with large quantities of reagent in order to enhance the sensitivity of the sensor. Physical entrapment onto the probe can also be another form of immobilization that is suitable to chemical or biological reagents. Iramobilization on cellulose or poly(vinyl chloride) films allows greater loading, but decreases response time because the reagent is immobilized in single layers.
Sensors for the detection of potassium ions are taught in U.S. Pat. No. 5,154,890 issued Oct. 15, 1992 to Mauze et al wherein 2,2-bis(3,4-(15-crown-5)-2-nitrophenylcarbamoxymethyl)tetradecanol-14 has at least one binding site and is provided with a fluorophore such as Rhodamine-B at that binding site. The sensing material is immobilized in a gel of polyacrylamide. U.S. Pat. No. 5,176,882 issued Jan. 5, 1993 to Gray et al teaches a dual fiberoptic cell for multiple serum measurements where both a gas and an ion are analyzed simultaneously using a single probe having two separate fiber optic sensors incorporated therein. The gas is detected by the color change of a dye and the ion is detected by the fluorescing of a fluorescent metal ion sensitive dye. While this reference mentions detecting calcium the only teaching and examples are restricted to metal ions of potassium, sodium and lithium. U.S. Pat. No. 5,093,266 issued Mar. 3, 1992 to Leader et al teaches the use of a sensor probe for the determining of ptI or carbon dioxide concentration of a liquid medium using a pH-sensitive fluorescent indicator 8-hydroxy-l,3,6-pyrenetrisulfonic acid (HPTA). Another pH sensor is disclosed in U.S. Pat. No. 5,118,405 issued Jun. 2, 1992 to Kaneko et al. U.S. Pat. No. 4,851,195 issued Jul. 25, 1989 to Matthews et al teaches another carbon dioxide sensor. Oxygen sensors and their method of use are disclosed, for example, in U.S. Pat. No. 4,974,929 issued Dec. 4, 1990 to Curry., U.S. Pat. No. 5,190,729 issued Mar. 2, 1993 and U.S. Pat. No. 4,861,727 issued Aug. 29, 1989 both to Hauenstein et al. U.S. Pat. No. 5,194,393 issued Mar. 16, 1993 to Hugl et al teaches an optical biosensor using a LB film and U.S. Pat. No. 5,227,134 issued Jul. 13, 1993 to Janata teaches a liquid/membrane cell sensor.
Many fluorescent probes have been developed to increase the usefulness of chemical sensors for non-fluorescent analytes. Sensor devices generally use chemical reagents immobilized at the terminus of an optical fiber that react with or respond to the presence of the analyte of interest. One such nonfluorescing calcium chelator used as a basis for the synthesis of some fluorescent analogues is ethyleneglycol-bis(.beta.-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA). The first fluorescent calcium indicator was 2-{[2-[bis(carboxymethyl)amino]-5'-methylphenoxy]methyl}-6-methoxy-8-[bis( carboxymethyl)amino]quinoline (Quin-2), R. Y. Tsien, Biochemistry, 19, 2396 (1980). Calcium binding leads to an increase of fluorescence intensity of this compound, but its use in living cells is limited. Other types of fluorescent calcium indicators include 1H-indole-6-carboxylic acid,2-(4-bis-(carboxymethyl)amino)-3(2-(2-(bis-carboxymethyl)amino-5-meth ylphenoxy)ethoxy)phenyl(Indo-1).
It is clear that there is a need for a metal ion sensor, particularly a calcium sensor, for real-time monitoring of metal ion levels in situ in a variety of situations including in vivo biological analysis and remote process control analysis.